Article

Reduction of Intersex in a Wild Fish Population in Response to Major Municipal Wastewater Treatment Plant Upgrades

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Abstract

Intersex in fish downstream of municipal wastewater treatment plants (MWWTPs) is a global concern. Consistent high rates of intersex in male rainbow darter (Etheostoma caeruleum) have been reported for several years in the Grand River, in southern Ontario, Canada, in close proximity to two MWWTPs. The larger MWWTP (Kitchener) recently underwent upgrades that included the conversion from a carbonaceous activated sludge to nitrifying activated sludge treatment process. This created a unique opportunity to assess whether upgrades designed to improve effluent quality could also remediate the intersex previously observed in wild fish. Multiple years (2007-2012) of intersex data on male rainbow darter collected before the upgrades at sites associated with the MWWTP outfall were compared with intersex data collected in post-upgrade years (2013-2015). These upgrades resulted in a reduction from 70-100% intersex incidence (pre-upgrade) to <10% in post-upgrade years. Although the cause of intersex remains unknown, indicators of effluent quality including nutrients, pharmaceuticals, and estrogenicity improved in the effluent after the upgrades. This study demonstrated that investment in MWWTP upgrades improved effluent quality and was associated with an immediate change in biological responses in the receiving environment. This is an important finding considering the tremendous cost of wastewater infrastructure.

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... Impacts below the MWWTP outfalls have primarily been observed in males at multiple levels of biological organization. These effects include increased expression of vitellogenin (Bahamonde et al., 2014;Fuzzen et al., 2016), decreased steroid hormone production (e.g., 11-ketotestosterone) (Tetreault et al., 2011;Fuzzen et al., 2016), delayed sperm development (Fuzzen et al., 2016), and high incidences and severe cases of intersex (Tetreault et al., 2011;Tanna et al., 2013;Bahamonde et al., 2015b;Fuzzen et al., 2015;Fuzzen et al., 2016). For all these endpoints, intersex was the most consistent effect observed across multiple years and seasons (Fuzzen et al., 2016). ...
... Impacts below the MWWTP outfalls have primarily been observed in males at multiple levels of biological organization. These effects include increased expression of vitellogenin (Bahamonde et al., 2014;Fuzzen et al., 2016), decreased steroid hormone production (e.g., 11-ketotestosterone) (Tetreault et al., 2011;Fuzzen et al., 2016), delayed sperm development (Fuzzen et al., 2016), and high incidences and severe cases of intersex (Tetreault et al., 2011;Tanna et al., 2013;Bahamonde et al., 2015b;Fuzzen et al., 2015;Fuzzen et al., 2016). For all these endpoints, intersex was the most consistent effect observed across multiple years and seasons (Fuzzen et al., 2016). ...
... Impacts below the MWWTP outfalls have primarily been observed in males at multiple levels of biological organization. These effects include increased expression of vitellogenin (Bahamonde et al., 2014;Fuzzen et al., 2016), decreased steroid hormone production (e.g., 11-ketotestosterone) (Tetreault et al., 2011;Fuzzen et al., 2016), delayed sperm development (Fuzzen et al., 2016), and high incidences and severe cases of intersex (Tetreault et al., 2011;Tanna et al., 2013;Bahamonde et al., 2015b;Fuzzen et al., 2015;Fuzzen et al., 2016). For all these endpoints, intersex was the most consistent effect observed across multiple years and seasons (Fuzzen et al., 2016). ...
Thesis
Impacts on aquatic biota residing near municipal wastewater treatment plant (MWWTP) outfalls have been documented globally. These impacts may be directly or indirectly associated with elevated contaminants such as nutrients, metals, suspended solids (SS), biochemical oxygen demanding matter (BOD), pharmaceuticals, and personal care products. A variety of effects have been well documented in the Grand River watershed of southern Ontario below the outfalls of the MWWTPs of the cities of Kitchener and Waterloo. Responses in wild fish have been reported at multiple levels of biological organization, ranging from altered gene expression to changes in fish communities, as well as changes in nutrient cycling within the aquatic food web. The most consistently observed effect has been high occurrences and severe cases of intersex (ova-testes) in the male rainbow darter (Etheostoma Caeruleum); this finding represents one of the worst examples of pollution-caused intersex reported anywhere in the world. Primarily in response to the introduction of new effluent quality standards, the Region of Waterloo has invested millions of dollars to upgrade several of its MWWTPs including the facility servicing Kitchener, creating a unique opportunity to conduct a before-and-after study. The main objective of this thesis was to assess if treatment upgrades, which were targeted at conventional contaminants (i.e., ammonia, BOD, SS, and chloride), effectively remediated the responses previously reported in wild fish downstream of the MWWTP. To test this, historical, archived, and new data collections were used to assess changes at multiple levels of biological organization, including changes in nutrient cycling in the aquatic food web, reproductive effects in the male rainbow darter (e.g., intersex), and changes in fish community composition. For comparative purposes, responses in rainbow darter were also examined at numerous reference sites and below the smaller Waterloo MWWTP, which did not undergo any major upgrades during the study period. The treatment upgrades at the Kitchener MWWTP (which included nitrifying activated sludge) improved the overall quality of the effluent; these improvements included reductions in nutrients (total ammonia), pharmaceuticals, and total estrogenicity (E2eq). In contrast, the Waterloo MWWTP had deteriorating effluent quality, with ammonia levels increasing over the course of the study. Changes in effluent quality at both the Kitchener and Waterloo MWWTPs were detected in the downstream aquatic food webs using stable isotope ratios (δ15N and δ13C). Patterns of δ15N in a primary consumer (benthic invertebrate) and a secondary consumer (rainbow darter) reflected the exposure to MWWTP effluents and changes in nutrient cycling in response to the changing effluent quality. A major reduction in intersex in the male rainbow darter below the Kitchener MWWTP outfall was also associated with the improvements in effluent quality. Rates of intersex were reduced by as much as 70% in the first year post-upgrade and dropped to near background levels within three years. Detecting change in fish communities below MWWTP outfalls (including before and after the upgrades) was more challenging. While subtle changes were detected (e.g., increases in pollution-tolerant species below the MWWTP outfalls), these could not be directly associated with MWWTP effluents because they were confounded by a watershed gradient (e.g., stream size). Fish communities were highly variable both spatially and temporally, limiting our ability to associate changes with local environmental conditions (i.e., effects of MWWTP outfalls). Although rainbow darter has been used as a sentinel species for detecting impacts of MWWTP effluents in many studies, little is known about its movement patterns. Elevated intersex was observed historically at the near-field upstream site of the Kitchener MWWTP outfall, leading to a hypothesis that wastewater-exposed fish may be moving upstream. To inform the interpretation of responses in rainbow darter as a sentinel species, a mark-and-recapture study was conducted at an upstream reference site to better understand their movement. Although the majority of fish (85%) had high site fidelity, a small proportion of fish moved considerable distances (up to 975 m). This study confirmed that there is potential for some fish to move and thereby confound the interpretation of near-field upstream sites that are not physically separated from the sites below the MWWTP outfall. The decline in intersex in rainbow darter after the upgrades at the site immediately upstream of the Kitchener outfall supports the view that at least some of the responses seen at this site were probably associated with fish movements. Overall, this thesis advances our understanding of the impacts of MWWTP effluents on wild fish and their response to improved effluent quality (i.e., treatment). The relatively simple (conventional) upgrades at the Kitchener MWWTP resulted in improvements in the aquatic receiving environment, indicating that more advanced treatment may not be required to address these effects of concern. However, other impacts may be occurring that were not measured in this study. The results drawn from this thesis may have implications for future wastewater management strategies for other MWWTPs across Canada and around the globe. In addition, these studies may provide insight into key biological endpoints that could be useful for future biomonitoring programs for MWWTP effluents.
... The watershed has been the subject of many studies that evaluated the responses of rainbow darters (Etheostoma caeruleum) to municipal wastewater effluent exposures. Changes in gene expression, stress response, male fish feminization (intersex), and fish community assemblages have been reported (Bahamonde et al., 2014;Fuzzen et al., 2016;Hicks et al., 2017a;Hicks et al., 2017b;Ings et al., 2012;Marjan et al., 2017;Tanna et al., 2013;Tetreault et al., 2011). The severity of these biological responses is typically higher downstream of the two major WWTPs (Kitchener and Waterloo WWTPs; Fig. 1). ...
... The model predictions were assessed using a total estrogenicity dataset (E2 equivalence) that was derived from the YES assay conducted at various times over the period of 2009 to 2015 (methodology described in Hicks et al., 2017a). There are other estrogens present in the municipal wastewater effluent such as bisphenol A and nonylphenols that can contribute to the total estrogenicity. ...
... These results suggest that the operation of the partially upgraded Kitchener WWTP is not expected to impair reproductive health of fish that might be exposed to its effluent. This finding supports the work of Hicks et al. (2017a) that indicate significant reduction of fish intersex incidence and severity after the upgrades of the Kitchener WWTP. ...
Article
In this study, the estrogenicity of two major wastewater treatment plant (WWTP) effluents located in the central reaches of the Grand River watershed in southern Ontario was estimated using population demographics, excretion rates, and treatment plant-specific removals. Due to the lack of data on estrogen concentrations from direct measurements at WWTPs, the treatment efficiencies through the plants were estimated using the information obtained from an effects-directed analysis. The results show that this approach could effectively estimate the estrogenicity of WWTP effluents, both before and after major infrastructure upgrades were made at the Kitchener WWTP. The model was then applied to several possible future scenarios including population growth and river low flow conditions. The scenario analyses showed that post-upgrade operation of the Kitchener WWTP will not release highly estrogenic effluent under the 2041 projected population increase (36%) or summer low flows. Similarly, the Waterloo WWTP treatment operation is also expected to improve once the upgrades have been fully implemented and is expected to effectively treat estrogens even under extreme scenarios of population growth and river flows. The developed model may be employed to support decision making on wastewater management strategies designed for environmental protection, especially on reducing the endocrine effects in fish exposed to WWTP effluents.
... The rainbow darter (Etheostoma caeruleum), a small-bodied species (up to 80 mm), has recently been used as a sentinel species in several biomonitoring studies to assess the effects of municipal wastewater in the Grand River watershed in southern Ontario (Fuzzen et al., 2016;Hicks, Fuzzen, et al., 2017a;Hicks, Loomer, et al., 2017b). This benthic species is short lived, reaching a maximum age of 5 years (Beckman, 2002), are sexually mature at age 1+ and spawn in the spring (Winn, 1958). ...
... In addition, sewage outfalls decrease water quality, and this may result in unfavourable conditions (e.g., low dissolved oxygen) that cause fish to emigrate (Lucas & Baras, 2001). Before 2012, the Kitchener WWTP effluent resulted in low oxygen and elevated ammonia concentrations downstream of the outfall (Hicks, Fuzzen, et al., 2017a). However, 3 years after process upgrades were implemented at the Kitchener WWTP, no highly intersex fish in the upstream (or downstream) site were recorded (Hicks, Fuzzen, et al., 2017a), suggesting that before the upgrades highly intersex fish (likely exposed to sewage) were probably moving upstream. ...
... Before 2012, the Kitchener WWTP effluent resulted in low oxygen and elevated ammonia concentrations downstream of the outfall (Hicks, Fuzzen, et al., 2017a). However, 3 years after process upgrades were implemented at the Kitchener WWTP, no highly intersex fish in the upstream (or downstream) site were recorded (Hicks, Fuzzen, et al., 2017a), suggesting that before the upgrades highly intersex fish (likely exposed to sewage) were probably moving upstream. The results of this study suggest that although most rainbow darters remain within a small home range, there is potential for some fish to move and confound the interpretation of near-field upstream sites that are not physically separated. ...
Article
Small‐bodied fish species are commonly used for the assessment of environmental effects because they are short lived, abundant, and they mature early. Although they are generally considered to be less mobile than larger bodied species, relatively little is known about their movement patterns. In this study, we tagged 3,001 rainbow darters (Etheostoma caeruleum) (≤76 mm) in the upper Grand River of southern Ontario with visible implant alpha tags and elastomers in 3 riffles. Five hundred sixty‐five fish were recaptured over 4 recapture events (including spawning and nonspawning periods) over a spatial extent of 1900 m. The rainbow darter demonstrated high site fidelity having a median movement of 5 m and with 85% staying within the riffle in which they were originally tagged. Most movements occurred during the spawning period, where males moved at a greater frequency and had a tendency to move longer distances (up to 975 m). There was also a bias in the direction of movement, which was dependent on the recapture season. Overall, the high site fidelity of the rainbow darter makes it a candidate, sentinel species for the assessment of environmental effects.
... Ten sites along the central Grand River spanning 60 km were sampled for fish in October 22-27, 2018 (Fig. 1); the sites were chosen based on their location relative to two large municipal secondary-conventional activated sludge treatment plants, the Waterloo and Kitchener WWTPs (Table 1, Fig. 1), that have been described in previous studies (Mehdi et al., 2017;Tetreault, 2012;Fuzzen et al., 2015;Fuzzen, 2016;Tetreault et al., 2013;Srikanthan, 2019). The Waterloo and Kitchener WWTPs serve approximately 98,000 and 219,000 people, respectively (Arlos et al., 2018b); upgrades have occurred recently at both plants and information on their operations are described in Hicks et al. (2017b). Our study is part of a larger program studying the impacts of wastewater effluents on rainbow darter (Tetreault, 2012;Tetreault et al., 2013;Fuzzen et al., 2015;Fuzzen, 2016;Hicks et al., 2017a;Hicks et al., 2017b). ...
... The Waterloo and Kitchener WWTPs serve approximately 98,000 and 219,000 people, respectively (Arlos et al., 2018b); upgrades have occurred recently at both plants and information on their operations are described in Hicks et al. (2017b). Our study is part of a larger program studying the impacts of wastewater effluents on rainbow darter (Tetreault, 2012;Tetreault et al., 2013;Fuzzen et al., 2015;Fuzzen, 2016;Hicks et al., 2017a;Hicks et al., 2017b). Several studies have characterized the chemical influences and water flow (Arlos et al., 2014, Arlos et al., 2018a, Arlos et al., 2018b, Srikanthan et al., 2019 at sites upstream and downstream of the Waterloo and Kitchener WWTPs. ...
... Several studies have characterized the chemical influences and water flow (Arlos et al., 2014, Arlos et al., 2018a, Arlos et al., 2018b, Srikanthan et al., 2019 at sites upstream and downstream of the Waterloo and Kitchener WWTPs. Furthermore, while there are numerous small WWTPs, agricultural inputs, and influences of large dams in the upper Grand River that alter nutrient cycling and aquatic community metabolism, the Waterloo and Kitchener WWTPs are major point source inputs of nutrients and contaminants (Loomer et al., 2015;Hicks et al., 2017a;Venkiteswaran et al., 2015), even after recent infrastructure upgrades (Hicks et al., 2017b). ...
Article
Municipal wastewater treatment plant (WWTP) effluent contains pharmaceuticals and personal care products known to affect fish health and reproduction. The microbiome is a community of bacteria integral in maintaining host health and is influenced by species, diet, and environment. This study investigated changes in the diversity and composition of the gut content microbiome of rainbow darter (Etheostoma caeruleum) at ten sites on the Grand River, Ontario, Canada. Gut contents were collected in Fall 2018 from rainbow darter at sites upstream and downstream of two municipal wastewater treatment plants (WWTPs; Waterloo and Kitchener). 16S rRNA genes were sequenced to determine the composition and diversity (alpha and beta) of microbial taxa present. Gut content bacterial diversity increased downstream of both WWTP outfalls; dominance of bacterial Amplicon Sequence Variants (ASVs) decreased compared to upstream fish. Fish collected at different sites had distinct bacterial communities, with upstream samples dominant in Proteobacteria and Firmicutes, and downstream samples increasingly abundant in Proteobacteria and Cyanobacteria. In mammals, increased abundance of Proteobacteria is indicative of microbial dysbiosis and has been linked to altered health outcomes, but this is not yet known for fish. This research indicates that the fish gut content microbiome was altered downstream of WWTP effluent outfalls and could lead to negative health outcomes.
... Tissues, organs and systems are involved in vertebrate toxicokinetics: integument, respiratory and digestive organs are firstly subjected to pollutant exposure, due to their direct interface with the outside (Fatima et al., 2014;Alves et al., 2016;Salamat and Zarie, 2016;Strzyzewska et al., 2016); besides, nervous, immune, and endocrine systems are mostly studied (Kumari and Khare, 2018;Lonappan et al., 2016;Vogt et al., 2018;Xu et al., 2018;Gambardella et al., 2016;Adeogun et al., 2016;Hicks et al., 2017;Tulloch et al., 2016a). ...
... Moreover, several Authors (e.g. Koh et al., 2009;McAdam et al., 2010;Verlicchi et al., 2012;Hicks et al., 2017) evidenced the positive effect of an efficient nitrification on EDCs removal. ...
Article
This opinion paper focuses on the role of eco-toxicological tools in the assessment of possible impacts of emerging contaminants on the aquatic ecosystem, hence, on human health. Indeed, organic trace pollutants present in raw and treated wastewater are the pivot targets: a multidisciplinary approach allows defining the basic principles for managing this issue, from setting a proper monitoring campaign up to evaluating the optimal process treatment. Giving hints on trace pollutants fate and behaviour, attention is focused on the choice of the bioassay(s), by analysing the meaning of possible biological answers. Data interpretation and exploitation are detailed with the final goal of providing criteria in order to be able to select the best targeted treatment options. The manuscript deals with conventional and innovative analytical approaches for assessing toxicity, by reviewing laboratory and field assays; illustrative real scale and laboratory applications integrate and exemplify the proposed approach.
... Sources of endocrine disrupting compounds (EDCs) are widespread, but waste water treatment plants (WWTPs), pulp mills, and agricultural practices (e.g. crop and livestock production) are important, well studied sources of EDCs to the environment (Frye et al., 2012;Hicks et al., 2017;Pollock et al., 2010). Although EDCs have been documented extensively in natural systems, it is often unclear if their presence has demonstrable adverse effects to fish and wildlife (Mills and Chichester, 2005). ...
... That stated, in the current study perhaps the most notable observation was the continued decrease in the observation of intersex. Similar findings of decreased intersex prevalence were observed in rainbow darter (Etheostoma caeruleum) inhabiting the Grand River in Ontario, Canada following WWTP upgrades (Hicks et al., 2017). It is possible that perhaps unreported changes in land-use activity in the Missisquoi watershed have ameliorated the problem, and we observed recovery during the time of this investigation. ...
Article
A reconnaissance project completed in 2009 identified intersex and elevated plasma vitellogenin in male smallmouth bass inhabiting the Missisquoi River, VT. In an attempt to identify the presence and seasonality of putative endocrine disrupting chemicals or other factors associated with these observations, a comprehensive reevaluation was conducted between September 2012 and June 2014. Here, we collected smallmouth bass from three physically partitioned reaches along the river to measure biomarkers of estrogenic endocrine disruption in smallmouth bass. In addition, polar organic chemical integrative samples (POCIS) were deployed to identify specific chemicals associated with biological observations. We did not observe biological differences across reaches indicating the absence of clear point source contributions to the observation of intersex. Interestingly, intersex prevalence and severity decreased in a stepwise manner over the timespan of the project. Intersex decreased from 92.8% to 28.1%. The only significant predictor of intersex prevalence was year of capture, based on logistic regression analysis. The mixed model of fish length and year-of-capture best predicted intersex severity. Intersex severity was also significantly different across late summer and early spring collections indicating seasonal changes in this metric. Plasma vitellogenin and liver vitellogenin Aa transcript abundance in males did not indicate exposure to estrogenic endocrine disrupting chemicals at any of the four sample collections. Analysis of chemicals captured by the POCIS as well as results of screening discrete water samples or POCIS extracts did not indicate the contribution of appreciable estrogenic chemicals. It is possible that unreported changes in land-use activity have ameliorated the problem, and our observations indicate recovery. Regardless, this work clearly emphasizes that single, snap shot sampling for intersex may not yield representative data given that the manifestation of this condition within a population can change dramatically over time.
... Similar results were described in studies performed by Jobling et al., [64] where estrogens (E1, E2, and EE2) have impacted population of roach (Rutilus rutilus) in British rivers. Results presented by Hicks et al. [65] where upgrades to the municipal wastewater treatment plant (Grand River, Canada) and as an effect lower the amount of estrogens in environment have led to rapid decline of intersex rainbow darter (Etheostoma caeruleum) males strongly correspond with those presented by Kidd and Jobling. On the other hand, in studies performed by Wang et al. [66] EE2 concentrations detected in Liaodong Bay (China) 0.42 ng/L, were too low to affect Wild Soiuy Mullets (Mugil soiuy) population. ...
Article
Full-text available
Nowadays, there is a growing interest in environmental pollution; however, knowledge about this aspect is growing at an insufficient pace. There are many potential sources of environmental con-tamination, including sex hormones—especially estrogens. The analyzed literature shows that es-trone (E1), estradiol (E2), estriol (E3), and synthetic ethinyloestradiol (EE2) are the most significant in terms of environmental impact. Potential sources of contamination are, among others, livestock farms, slaughterhouses, and large urban agglomerations. Estrogens occurring in the environment can negatively affect the organisms, such as animals, through phenomena such as feminization, dysregulation of natural processes related to reproduction, lowering the physiological condition of the organisms, disturbances in the regulation of both proapoptotic and anti-apoptotic processes, and even the occurrence of neoplastic processes thus drastically decreasing animal welfare. Un-fortunately, the amount of research conducted on the negative consequences of their impact on animal organisms is many times smaller than that of humans, despite the great richness and di-versity of the fauna. Therefore, there is a need for further research to help fill the gaps in our knowledge.
... Rainbow darter (Etheostoma caeruleum) are a small-bodied, freshwater, benthic and carnivorous fish found in northeastern North America, and known to have a small home range of approximately 5.0 m (Hicks et al., 2017b), making them valuable as a sentinel species in field studies. In particular, this species has been used for field studies of WWTP effluents (Fuzzen, 2016;Fuzzen et al., 2015;Hicks et al., 2017a;Mehdi et al., 2017;Robinson et al., 2016;Tetreault et al., 2012Tetreault et al., , 2014, including one study of their gut microbiome (Restivo et al., 2021). The rainbow darter is amenable to laboratory rearing and holding (Fuzzen, 2016;Dhakal, 2017;Mehdi et al., 2017;Turner and Bucking, 2019) and has been used to examine the effects of diet on the gut microbiome (Dhakal, 2017, Turner andBucking, 2019), making them ideal for the present study. ...
Article
An increasing number of laboratory studies are showing that environmental stressors and diet affect the fish gut microbiome. However, the application of these results to wild populations is uncertain as little is known about how the gut microbiome shifts when fish are transitioned from the field to the laboratory. To assess this, intestinal contents (i.e. digesta) of wild-caught rainbow darter (Etheostoma caeruleum) were sampled in the field and in the lab after 14- and 42-days acclimation. In addition, from days 15–42 some fish were exposed to waterborne triclosan, an antimicrobial found in aquatic ecosystems, or to dilutions of municipal wastewater effluents, to determine how these stressors affect the bacterial communities of gut contents. 16S rRNA gene amplicon sequencing was used to determine microbial community composition, alpha, and beta diversity present in the fish gut contents. In total, there was 8,074,658 reads and 11,853 amplicon sequence variants (ASVs) identified. The gut contents of wild fish were dominant in both Proteobacteria (35%) and Firmicutes (27%), while lab fish were dominant in Firmicutes (37–47%) and had lower alpha diversity. Wild fish had greater ASVs per sample (423-1304) compared to lab fish (19-685). Similarly, the beta-diversity of these bacterial communities differed between field and lab control fish; control fish were distinct from the 10% wastewater effluent and 100 ng/L TCS treatment groups. Results indicate that the gut microbiome of wild fish changes with the transition to laboratory environments; hence, prolonged acclimation to new settings may be required to achieve a stable gut content microbiome in wild-caught fish. Research is required to understand the length of time required to reach a stable fish gut microbiome.
... Where technological improvements of sewage treatment have been implemented, there have been reductions in intersex (feminised) fish caused by steroid oestrogens and their mimics, as well as improvements in river biodiversity. 40,41 These improvements, however, are costly. Owen and Jobling (2012) 42 estimated that upgrading all the WWTPs in England and Wales to comply with EU regulation to bring synthetic oestrogens below an average of 0.035 pg/L ethinylestradiol per annum would cost an estimated £26 billion. ...
Article
Full-text available
The influence of pharmaceuticals on the environment is an increasing concern among environmental toxicologists. It is known that their growing use is leading to detectable levels in wastewater, conceivably causing harm to aquatic ecosystems. Psychotropic medication is one such group of substances, particularly affecting high-income countries. While these drugs have a clear place in therapy, there is debate around the risk/benefit ratio in patients with mild mental health problems. Therefore, it is necessary to evaluate the wider implications as risks could extend beyond the individual to non-target organisms, particularly those in rivers and estuaries. Declaration of interest None.
... A key response also noted in the male darters to municipal effluent exposure was the induction VTG. Recent upgrades in sewage treatment within the watershed have resulted in reduced prevalence of intersex in the darters (Hicks et al., 2017). In other studies, upgrades of the wastewater treatment plant in Boulder Colorado, resulted in improved removal efficiency for many EDCs, particularly 17β-estradiol and estrone, and fathead minnow exposed to the post upgrade effluent showed reduced endocrine disruption relative to pre-upgrade conditions (Barber et al., 2012). ...
Article
Endocrine disrupting chemicals (EDCs) are ubiquitous in aquatic and terrestrial environments. The main objective of this review was to summarize the current knowledge of the impacts of EDCs on reproductive success in wildlife and humans. The examples selected often include a retrospective assessment of the knowledge of reproductive impacts over time to discern how the effects of EDCs have changed over the last several decades. Collectively, the evidence summarized here within reinforce the concept that reproduction in wildlife and humans is negatively impacted by anthropogenic chemicals, with several altering endocrine system function. These observations of chemicals interfering with different aspects of the reproductive endocrine axis are particularly pronounced for aquatic species and are often corroborated by laboratory-based experiments (i.e. fish, amphibians, birds). Noteworthy, many of these same indicators are also observed in epidemiological studies in mammalian wildlife and humans. Given the vast array of reproductive strategies used by animals, it is perhaps not surprising that no single disrupted target is predictive of reproductive effects. Nevertheless, there are some general features of the endocrine control of reproduction, and in particular, the critical role that steroid hormones play in these processes that confer a high degree of susceptibility to environmental chemicals. New research is needed on the implications of chemical exposures during development and the potential for long-term reproductive effects. Future emphasis on field-based observations that can form the basis of more deliberate, extensive, and long-term population level studies to monitor contaminant effects, including adverse effects on the endocrine system, are key to addressing these knowledge gaps.
... Although outfall concentrations are sometimes reduced (e.g. acetaminophen, oestrogens) by more advanced treatment processes -and with subsequent benefits downstream (Hicks et al., 2017) -some emerging contaminants (carbazepine, triclosan and diclofenac) are more recalcitrant and require the development of novel interventions (e.g. Bean et al., 2016). ...
Article
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In the 12 years since Dudgeon et al. (2006) reviewed major pressures on freshwater ecosystems, the biodiversity crisis in the world's lakes, reservoirs, rivers, streams and wetlands has deepened. While lakes, reservoirs and rivers cover only 2.3% of the Earth's surface, these ecosystems host at least 9.5% of the Earth's described animal species. Furthermore, using the World Wide Fund for Nature's Living Planet Index, freshwater population declines (83% between 1970 and 2014) continue to outpace contemporaneous declines in marine or terrestrial systems. The Anthropocene has brought multiple new and varied threats that disproportionately impact freshwater systems. We document 12 emerging threats to freshwater biodiversity that are either entirely new since 2006 or have since intensified: (i) changing climates; (ii) e‐commerce and invasions; (iii) infectious diseases; (iv) harmful algal blooms; (v) expanding hydropower; (vi) emerging contaminants; (vii) engineered nanomaterials; (viii) microplastic pollution; (ix) light and noise; (x) freshwater salinisation; (xi) declining calcium; and (xii) cumulative stressors. Effects are evidenced for amphibians, fishes, invertebrates, microbes, plants, turtles and waterbirds, with potential for ecosystem‐level changes through bottom‐up and top‐down processes. In our highly uncertain future, the net effects of these threats raise serious concerns for freshwater ecosystems. However, we also highlight opportunities for conservation gains as a result of novel management tools (e.g. environmental flows, environmental DNA) and specific conservation‐oriented actions (e.g. dam removal, habitat protection policies, managed relocation of species) that have been met with varying levels of success. Moving forward, we advocate hybrid approaches that manage fresh waters as crucial ecosystems for human life support as well as essential hotspots of biodiversity and ecological function. Efforts to reverse global trends in freshwater degradation now depend on bridging an immense gap between the aspirations of conservation biologists and the accelerating rate of species endangerment.
... Conventional sewage treatment has been reported to reduce EDCs significantly from high levels, for example 50 ng l −1 to below 0.75 ng l −1 (Leusch et al. 2005) and with EDCs removal efficiencies ranging from 70% to 100% (Kirk et al. 2002). Moreover, improved wastewater treatment in some cities has resulted in improved effluent quality and reduced effects of EDCs in the environment (Hicks et al. 2017). However, Bulawayo, and many other cities in developing countries, are characterised by aging and dysfunctional STP infrastructure. ...
Article
Peri-urban water bodies are at risk from excessive pollution as they are direct sinks for urban effluents. The occurrence of oestrogenic and androgenic endocrine disrupting chemicals (EDCs) in effluents and water bodies around the city of Bulawayo, Zimbabwe, and their effects on wild fish was investigated in 2013. Effluent and water were sampled from sewage treatment plants, an urban stream, and effluent-polluted dams, and were compared with a ‘pristine’ dam upstream of Bulawayo. Organic pollutants were extracted by solid-phase extraction and tested for EDCs using a yeast oestrogen/androgen screen. Oestrogenic and androgenic potencies were expressed as 17β-oestradiol equivalent (EEq) or dihydrotestosterone equivalent (TEq). Tilapia and catfish from the dams were analysed for gonado-somatic indices and testis histopathology. Effluents from STPs, which directly flow into Umguza Dam, had EEq of 33 ng l⁻¹ and 55 ng l⁻¹, respectively. Umguza Dam, Khami Dam and Matsheumhlope Stream had EEqs of 237 ng l–1, 9 ng l⁻¹ and 2 ng l⁻¹, respectively. Androgenic activity was detected in only one STP (TEq = 93 ng l⁻¹). Tilapia sampled from effluent-polluted dams had high incidences of testis-ova, but catfish had no signs of reproductive dysfunction. These findings underscore the need for greater attention to EDCs in developing countries where there is scant literature regarding their occurrence and impacts.
... Plants OX, MH, HG, and E were operated to nitrify (Table 2) and showed high removal of TCS (Fig. 3). Removal of trace contaminants has been correlated with degree of nitrification in other studies (Batt et al. 2006;Hicks et al. 2017;Servos et al. 2005;Smyth et al. 2008). Lozano and coworkers reported high removal of TCS during nitrificationdenitrification processes (Lozano et al. 2013). ...
Article
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Triclosan (TCS) is an antimicrobial agent used in many personal care and cleaning products. It has been detected in most environmental compartments and the main entry pathway is wastewater effluents and biosolids. TCS was analyzed in 300 samples of raw influent, final effluent, and biosolids from 13 wastewater treatment plants (WWTPs) across Canada representing five types of typical wastewater treatment systems. TCS was almost always detected in influent (median 1480 ng/L), effluent (median 107 ng/L), and biosolids (median 8000 ng/g dry weight) samples. Removals of TCS from lagoons as well as secondary and advanced treatment facilities were significantly higher than primary treatment facilities (p < 0.001). TCS removal was strongly correlated with organic nitrogen removal. TCS removals at most lagoons and plants that use biological treatment were higher during summer compared with winter. However, no seasonal or temperature effects were observed at the two primary facilities, likely due to the absence of biological activity. Aerobically digested solids contained the lowest levels (median 555 ng/g) while anaerobically digested primary solids contained the highest levels of TCS (median 22,700 ng/g). The results of this large comprehensive study demonstrate that TCS is consistently present in wastewater and biosolids at relatively high concentrations and that removal from wastewater and levels in biosolids are strongly influenced by the wastewater and solids treatment types.
... This resulted in improved effluent quality with regard to nutrients, pharmaceuticals, and estrogenicity and, astonishingly, a reduction to less than 10% intersex postupgrade. 147 Perinatal Di-(2-ethylhexyl) Phthalate Exacerbates Anaphylaxis in Male Mouse Offspring, by Emily Mackey This work, which was conducted at the laboratory of Dr Adam Moeser at Michigan State University, stems from the discovery of a sexually dimorphic mast cell phenotype and associated disease pathophysiology. 148 Previously published data have demonstrated that adult female mice exhibited greater serum histamine and more severe mast cell-associated pathophysiology in response to immunoglobulin E (IgE)-mediated anaphylaxis and restraint stress models of mast cell activation, compared with males. ...
Article
During the past 20 years, investigations involving endocrine active substances (EAS) and reproductive toxicity have dominated the landscape of ecotoxicological research. This has occurred in concert with heightened awareness in the scientific community, general public, and governmental entities of the potential consequences of chemical perturbation in humans and wildlife. The exponential growth of experimentation in this field is fueled by our expanding knowledge into the complex nature of endocrine systems and the intricacy of their interactions with xenobiotic agents. Complicating factors include the ever-increasing number of novel receptors and alternate mechanistic pathways that have come to light, effects of chemical mixtures in the environment versus those of single EAS laboratory exposures, the challenge of differentiating endocrine disruption from direct cytotoxicity, and the potential for transgenerational effects. Although initially concerned with EAS effects chiefly in the thyroid glands and reproductive organs, it is now recognized that anthropomorphic substances may also adversely affect the nervous and immune systems via hormonal mechanisms and play substantial roles in metabolic diseases, such as type 2 diabetes and obesity.
... Adverse effects have already been identified and continue to be related to the presence of pollutants in water, such as inhibition of neurotransmitters (de Oliveira et al., 2016), mutagenicity (Lutterbeck et al., 2015), carcinogenicity, biomagnification in marine trophic nets (Xue et al., 2017), phytotoxicity (Richter et al., 2016), feminization of fish (Hicks et al., 2017) and development of bacteria resistant to antibiotics (Miranda et al., 2016). ...
Article
Persistent organic products are compounds used for various purposes, such as personal care products, surfac-tants, colorants, industrial additives, food, pesticides and pharmaceuticals. These substances are constantly introduced into the environment and many of these pollutants are difficult to degrade. Toxic compounds classified as MoA 1 (Mode of Action 1) are low toxicity compounds that comprise nonreactive chemicals. In silico methods such as Quantitative Structure-Activity Relationships (QSARs) have been used to develop important models for prediction in several areas of science, as well as aquatic toxicity studies. The aim of the present study was to build a QSAR model-based set of theoretical Volsurf molecular descriptors using the fish acute toxicity values of compounds defined as MoA 1 to identify the molecular properties related to this mechanism. The selected Partial Least Squares (PLS) results based on the values of cross-validation coefficients of determination (Q cv 2) show the following values: Q cv 2 = 0.793, coefficient of determination (R 2) = 0.823, explained variance in external prediction (Q ext 2) = 0.87. From the selected descriptors, not only the hydrophobicity is related to the toxicity as already mentioned in previously published studies but other physicochemical properties combined contribute to the activity of these compounds. The symmetric distribution of the hydrophobic moieties in the structure of the compounds as well as the shape, as branched chains, are important features that are related to the toxicity. This information from the model can be useful in predicting so as to minimize the toxicity of organic compounds.
... Urban land use, the number of surface water discharge permits (including wastewater discharges) and the number of known contaminated sites were all positively correlated to estrogenicity in our study. Although we did not have enough sites to statistically compare intersex data to known point sources, similar to other studies [51,52], the presence of these point sources could have contributed to intersex observed across our study area. However, for several of the impoundments (Echo Lake, Canistear Reservoir and Splitrock Reservoir), there are no well-defined point sources that would contribute endocrine active chemicals into the impoundments where fish were sampled during the New Jersey reconnaissance study. ...
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The observation of testicular oocytes in male fishes has been utilized as a biomarker of estrogenic endocrine disruption. A reconnaissance project led in the Northeastern United States (US) during the period of 2008–2010 identified a high prevalence of intersex smallmouth bass on or near US Fish & Wildlife Service National Wildlife Refuges that included the observation of 100% prevalence in smallmouth bass males collected from the Wallkill River, NJ, USA. To better assess the prevalence of intersex smallmouth bass across the state of New Jersey, a tiered reconnaissance approach was initiated during the fall of 2016. Surface water samples were collected from 101 (85 river, 16 lake/reservoir) sites across the state at base-flow conditions for estrogenicity bioassay screening. Detectable estrogenicity was observed at 90% of the sites and 64% were above the US Environmental Protection Agency trigger level of 1 ng/L. Median surface water estrogenicity was 1.8 ng/L and a maximum of 6.9 ng/L E2EqBLYES was observed. Adult smallmouth bass were collected from nine sites, pre-spawn during the spring of 2017. Intersex was identified in fish at all sites, and the composite intersex prevalence was 93.8%. Prevalence across sites ranged from 70.6% to 100%. In addition to intersex, there was detectable plasma vitellogenin in males at all sites. Total estrogenicity in surface water was determined at these fish collection sites, and notable change over time was observed. Correlation analysis indicated significant positive correlations between land use (altered land; urban + agriculture) and surface water estrogenicity. There were no clear associations between land use and organismal metrics of estrogenic endocrine disruption (intersex or vitellogenin). This work establishes a baseline prevalence of intersex in male smallmouth bass in the state of New Jersey at a limited number of locations and identifies a number of waterbodies with estrogenic activity above an effects-based threshold.
... The environmental occurrence of BCs downstream from WWTP outfalls have been shown to adversely affect fish health in several studies (Vajda et al., 2008;Corcoran et al., 2010;Barber et al., 2011;Jorgenson et al., 2018;Meador et al., 2018), and mixtures of BCs may be considerably more toxic or active than the individual components of the mixture (Hayes et al., 2006;Wolfe et al., 2015;Heys et al., 2016;Parrish et al., 2019). In other cases, enhancements in treatment plant operations have been shown to improve conditions for fish Völker et al., 2016;Hicks et al., 2017;Marjan et al., 2017). Suspended sediments and plankton could also be accumulating BCs (Lahti and Oikari, 2011;Bai and Acharya, 2017;Fan et al., 2017) and hence be an additional source of exposure for Asian carp. ...
Article
Two non-native carp species have invaded the Illinois Waterway and are a threat to Great Lakes ecosystems. Poor water quality in the upper Illinois Waterway may be a factor contributing to the stalling of the carp population front near river mile 278. In 2015, the U.S. Geological Survey collected 4 sets of water samples from two sites upstream and 4 sites downstream from river mile 278, and one tributary. Each sample was analyzed for up to 649 unique constituents of which 287 were detected including 96 pesticides, 62 pharmaceuticals, 39 wastewater indicator chemicals, 29 metals, 19 volatile organic compounds (VOCs), 6 disinfection by-products (DBPs), 5 hormones, and 5 carboxylic acids. Potential for bioactivity was estimated by comparing chemical concentrations to aquatic life or human health criteria and to in-vitro bioactivity screening results in the U.S Environmental Protection Agency ToxCast™ database. The resulting hazard quotients and exposure-activity ratios (EARs) are toxicity indexes that can be used to rank potential bioactivity of individual chemicals and chemical mixtures. This analysis indicates that several bioactive chemicals (BCs) including: carbendazim, 2,4-D, metolachlor, terbuthylazine, and acetochlor (pesticides); 1,4-dioxane (VOC); metformin, diphenhydramine, sulfamethoxazole, tramadol, fexofenadine, and the anti-depressants (pharmaceuticals); bisphenol A, 4-nonylphenol, galaxolide, 4-tert-octylphenol (wastewater indicator chemical); lead and boron (metals); and estrone (hormone) all occur in the upper Illinois Waterway at concentrations that produce elevated EARs values and may be adversely affecting carp reproduction and health. The clear differences in water quality upstream and downstream from river mile 278 with higher contaminant concentrations and potential bioactivity upstream could represent a barrier to carp range expansion.
... 31 In 2015, 20% of the sewage sludge produced from MSTPs was applied to croplands, 31 thereby having the potential to increase the exposure of the Chinese population to Hg through rice and livestock (e.g., poultry) intake. 32,33 Few studies have focused on the release of MeHg from 34,35 The MeHg released from municipal sewage and other anthropogenic sources should not be ignored. ...
Article
As a globally transported pollutant, mercury (Hg) released from human activity and methylmercury (MeHg) in the food web are global concerns due to their increasing presence in the environment. In this study, we found that Hg released from municipal sewage into the environment in China is a substantial anthropogenic source based on mass sampling throughout China. In total, 160 Mg (140-190 Mg, from the 20th percentile to the 80th percentile) of Hg (THg) and 280 kg (240-330 kg) of MeHg were released from municipal sewage in China in 2015. The quantities of released THg and MeHg were the most concentrated in the coastal regions, especially in the East, North and South China regions. However, the per capita release of THg and MeHg was the highest in the Tibetan region, which is recognized as the cleanest region in China. THg released into aquatic environments was mitigated from 2001 to 2015 in China, but the amounts released into other sinks increased. This study provides the first picture of the release of Hg from municipal sewage into various sinks in China, and policy makers should pay more attention to the diversity and complexity of the sources and transport of Hg, which can lead to Hg accumulation in the food web and can threaten human health.
... Additionally, Cipoletti et al. (2019) reported delays in development and growth rate responses in larval and juvenile fathead minnows exposed to CECs present in the Maumee River. While the ecotoxic effects of many CECs and their potential impacts on ecosystems are poorly understood, there is growing evidence suggesting exposure to low concentrations of some of these chemicals and chemical mixtures have physiological effects on fish and other wildlife and can disrupt their endocrine and immune systems (Gonz alez et al., 2012;Jasinska et al., 2015;Hicks et al., 2017;Thomas et al., 2017;Cipoletti et al., 2019). In general these effects do not cause mortality and are not grossly observable in the individual animal, therefore they may be imperceptible until significant or catastrophic impairments to aquatic wildlife populations and communities are realized. ...
Article
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Recent research suggests contaminants of emerging concern (CECs) are widespread and environmentally relevant concentrations can impact fishes. However, little is known about impacts of CECs to long-lived or rare species. The objective of this study was to characterize CEC concentrations in lake sturgeon serum and gametes. Blood serum was collected non-lethally from lake sturgeon (Acipenser fulvescens) at four lower Great Lakes basin sites: Detroit, upper Niagara, lower Niagara, and St. Lawrence rivers; additionally, gametes were collected from lake sturgeon in the St. Lawrence River. Samples were analyzed for pharmaceuticals and personal care products (PPCPs) and polybrominated diphenyl ethers (PBDEs). Overall, 44 different PPCPs were identified in serum and gamete samples across sites, with 22 PPCPs identified in at least 25% of serum samples and three PPCPs identified in 25% of gamete samples. PPCP concentrations in serum and gametes ranged from 0.00208 to 130 ppb and 0.00538–190 ppb, respectively. NMDS ordination revealed differences in the presence and concentrations of PPCPs in lake sturgeon serum across sites, however, N,N-diethyl-meta-toluamide (DEET), hydrocortisone, benztropine, and amitriptyline were detected in at least one serum sample at all sites. Additionally, DEET, 10-hydroxy-amitriptyline, and sertraline were detected in ≥25% of gamete samples collected from the St. Lawrence River. Twenty-six PBDE congeners were identified in 25% of serum samples and 24 were identified in 25% of gamete samples. PBDEs in serum were present across all sites and in gametes of St. Lawrence River lake sturgeon, and total PBDE concentrations in serum and gametes ranged from 0.184 to 12.7 ppb and 0.0826–0.44 ppb, respectively. Managers of lake sturgeon populations may need to consider the impacts of CECs if reproductive, developmental, behavioral, growth effects, or mortality are observed in the Great Lakes basin or other areas that are impacted by increased exposures to PPCPs and PBDEs.
... To close this gap, several case studies were conducted during the last years which proved biological effect monitoring based on biotests and biomarkers to be a useful tool to assess the effluentrelated risk for aquatic organisms [11,12]. Moreover, comparison of data obtained prior and subsequent to the installation of a new wastewater treatment technology [13][14][15][16][17][18] or of samples taken at different steps within the treatment process [19,20] was shown to be a valuable strategy to evaluate the efficiency of this new technology. Especially in situ exposure (active monitoring) of caged fish has often been applied in this context [21,22]. ...
Article
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Background In situ exposure of rainbow trout up- and downstream of differently equipped wastewater treatment plants (WWTPs) and subsequent analyses of micronuclei frequencies and hepatic EROD activities were used to evaluate the impact of the effluents on fish health. Two of the facilities (WWTPs A and B) were conventional treatment plants. WWTP C has been equipped with a powdered activated carbon stage. Here, analyses were conducted prior and subsequent to this upgrade. Results Differences did not only occur when comparing conventional (WWTPs A, B and C prior to the upgrade) and advanced treatment (WWTP C after the upgrade), but also between the conventionally equipped WWTPs. There was no indication for genotoxic effects or pollution-related EROD induction in fish exposed at WWTP A. In contrast, trout exposed at WWTP B expressed strong reactions. However, here, adverse reactions were also observed in fish kept upstream. Similar observations were made for EROD activities in fish exposed at WWTP C prior to the upgrade, whereas genotoxic effects could only be seen in trout kept downstream of this effluent. Upgrading of WWTP C resulted in a significant reduction of both genotoxic effects and EROD levels. Conclusions The results show financial investments in advanced wastewater treatment to be beneficial for aquatic ecosystems, especially when conventional technologies do not sufficiently remove pollutants. Yet, negative impacts of effluents on aquatic organisms can, under certain conditions, also be avoided by conventional treatment. Therefore, we recommend deciding on the necessity and the type of WWTP upgrading on a case-by-case basis.
... The synthetic hormone 17αethinylestradiol was demonstrated to have caused the collapse of a fish population at concentrations detected in the environment (Kidd et al. 2007). Hicks et al. (2017) reported decreased incidence of intersex in a population of rainbow darters downstream of a municipal sewage treatment facility, after plant upgrades, with corresponding decreases in the concentration of estrogens detected in the effluent. However, observed effects in invertebrates have not been as clear-cut, and questions remain as to whether these compounds have any hormonal action in molluscs (Scott 2012(Scott , 2013. ...
Article
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The toxicity of endocrinologically active pharmaceuticals finasteride (FIN) and melengestrol acetate (MGA) was assessed in freshwater mussels, including acute (48 h) aqueous tests with glochidia from Lampsilis siliquoidea, sub-chronic (14 days) sediment tests with gravid female Lampsilis fasciola, and chronic (28 days) sediment tests with juvenile L. siliquoidea, and in chronic (42 days) sediment tests with the amphipod Hyalella azteca and the mayfly Hexagenia spp. Finasteride was not toxic in acute aqueous tests with L. siliquoidea glochidia (up to 23 mg/L), whereas significant toxicity to survival and burial ability was detected in chronic sediment tests with juvenile L. siliquoidea (chronic value (ChV, the geometric mean of LOEC and NOEC) = 58 mg/kg (1 mg/L)). Amphipods (survival, growth, reproduction, and sex ratio) and mayflies (growth) were similarly sensitive (ChV = 58 mg/kg (1 mg/L)). Melengestrol acetate was acutely toxic to L. siliquoidea glochidia at 4 mg/L in aqueous tests; in sediment tests, mayflies were the most sensitive species, with significant growth effects observed at 37 mg/kg (0.25 mg/L) (ChV = 21 mg/kg (0.1 mg/L)). Exposure to sublethal concentrations of FIN and MGA had no effect on the (luring and filtering) behaviour of gravid L. fasciola, or the viability of their brooding glochidia. Based on the limited number of measured environmental concentrations of both chemicals, and their projected concentrations, no direct effects are expected by these compounds individually on the invertebrates tested. However, organisms are exposed to contaminant mixtures in the aquatic environment, and thus, the effects of FIN and MGA as components of these mixtures require further investigation.
... River water was collected to assess the estrogenicity of the SAR at baseflow conditions. The mean in vitro EEQs for August/September and November sampling events were consistent with EEQs determined from previous studies evaluating the estrogenicity of the SAR and other effluent-dominated river systems (Hicks et al., 2017;Song et al., 2020;Vajda et al., 2008;Xie et al., 2004). EEQs ranged from <1 to 2.57 ng/L in effluent collected from Prado in April 2003 (Xie et al., 2004), and between 0.02 ng/L and 3.4 ng/L in sites located upstream and downstream of an effluent outfall in Boulder Creek in September 2003, respectively (Vajda et al., 2008). ...
Article
Estrogenically active compounds (EACs) in surface waters can disrupt the endocrine system of biota, raising concern for aquatic species. Concentrations of EACs are generally higher in effluent-dominated aquatic systems, such as California's Santa Ana River (SAR). Addressing estrogenicity of effluent-dominated waters is increasingly important due to both increasing urbanization and climate change. To this end, water samples were collected from multiple sites downstream of wastewater treatment plants (WWTPs) and intermittent points along the SAR during 2018–2019 and cell-based bioassays were used to determine estrogen receptor activity. During baseflow conditions, the highest estradiol equivalencies (EEQs) from all SAR water between summer (August and September) and fall (November) sampling events in 2018 were from Yorba Linda (EEQ = 1.36 ± 0.38 ng/L) and Prado (1.14 ± 0.13 ng/L), respectively. Water extracts in January 2019 following a major rainfall generally had higher EEQs with the highest EEQ of 10.0 ± 0.69 ng/L observed at Yorba Linda. During low flow conditions in November 2018, male Japanese medaka (Oryzias latipes) fish were exposed to SAR water to compare to cell bioassay responses and targeted analytical chemistry for 5 steroidal estrogens. Chemical-based EEQ correlations with in vitro EEQs were statistically significant. However, vitellogenin (vtg) mRNA expression in the livers of medaka exposed to SAR water was not significantly different compared to controls. These results indicate that seasonal variation and surface water runoff events influence estrogenic activity in the SAR and may induce estrogenic effects to native fish populations in wastewater-dominated streams in general.
... In past studies, effects on reproduction, growth, sex ratios, metabolism, and molecular signatures have all been identified as maladaptive phenotypes in downstream locations from MWWTPs in this area, which were attributed to effluent exposure [3][4][5]. Rainbow darters downstream from the MWWTPs in the Grand River have experienced changes in gene expression, skewed sex ratios, altered gonad size, and intersex or feminized fish populations are found, largely due to the presence of EDCs [6,7]. Additionally, run-off from urban developments and intense agricultural areas, along with stormwater drainages, are other sources of contamination and water quality degradation [8,9]. ...
Article
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The effluent from municipal wastewater treatment plants is a major point source of contamination in Canadian waterways. The improvement of effluent quality to reduce contaminants, such as pharmaceuticals and personal care products, before being released into the environment is necessary to reduce the impacts on organisms that live in the river downstream. Here, we aimed to characterize the metabolic and gill physiological responses of rainbow (Etheostoma caeruleum), fantail (Etheostoma flabellare), and greenside (Etheostoma blennioides) darters to the effluent in the Grand River from the recently upgraded Waterloo municipal wastewater treatment plant. The routine metabolism of darters was not affected by effluent exposure, but some species had increased maximum metabolic rates, leading to an increased aerobic scope. The rainbow darter aerobic scope increased by 2.2 times and the fantail darter aerobic scope increased by 2.7 times compared to the reference site. Gill samples from effluent-exposed rainbow darters and greenside darters showed evidence of more pathologies and variations in morphology. These results suggest that darters can metabolically adjust to effluent-contaminated water and may also be adapting to the urban and agricultural inputs. The modification and damage to the gills provide a useful water quality indicator but does not necessarily reflect how well acclimated the species is to the environment due to a lack of evidence of poor fish health.
... Our hypothesis was that the restored streams have lower levels of nutrients and EEDCs than the unrestored streams, although both restored and unrestored sub-watersheds have the potential to receive the same inputs. The effect of upgrades of WWTPs on EEDCs and nutrient reduction was evaluated by examining changes across wastewater treatment profiles of a WWTP (Barber et al., 2012;Hicks et al., 2017), as well as their upstream-downstream changes in chemical concentrations in receiving waters (Ciparis et al., 2012). Moreover, we compared nutrient and EEDCs in a CSO with those collected from WWTP effluents to assess potential effects of replacing aging CSO systems. ...
Article
We evaluate the impacts of different nutrient management strategies on the potential for co-managing estrogens and nutrients in environmental waters of the Potomac watershed of the Chesapeake Bay. These potential co-management approaches represent agricultural and urban runoff, wastewater treatment plant effluent, and combined sewer overflow replacements. Twelve estrogenic compounds and their metabolites were analysed by gas chromatography–mass spectrometry. Estrogenic activity (E2Eq) was measured by in vitro bioassay. We detected estrone E1 (0.05–6.97 ng L⁻¹) and estriol E3 (below detection-8.13 ng L⁻¹) and one conjugated estrogen (estrone-3-sulfate E1-3S; below detection-8.13 ng L⁻¹). E1 was widely distributed and positively correlated with E2Eq, water temperature, and dissolved organic carbon (DOC). Among nonpoint sources, E2Eq, and concentrations of E1, soluble reactive phosphorus (SRP) and total dissolved nitrogen (TDN) decreased by 51–61%, 77–82%, 62–64%, 4–16% in restored urban and agricultural streams with best management practices (BMPs) relative to unrestored streams without BMPs. In a wastewater treatment plant (Blue Plains WWTP), >94% of E1, E1-3S, E3, E2Eq and TDN were removed while SRP increased by 305% during nitrification/denitrification as a part of advanced wastewater treatment. Consequently, E1 and TDN concentrations in WWTP effluents were comparable or even lower than those observed in the receiving stream or river waters, and the effects of wastewater discharges on downstream E1 and TDN concentrations were minor. Highest E2Eq value and concentrations of E1, E3, and TDN were detected in combined sewer overflow (CSO). This study suggests that WWTP upgrades with biological nutrient removal, CSO management, and certain agricultural and urban BMPs for nutrient controls have the potential to remove estrogens from point and nonpoint sources along with other contaminants in streams and rivers.
... It is worth noting that environmental decision making is not always straightforward even when cost and political considerations are excluded. For example, enhanced WWTP N oxidation without denitrification may mitigate cHABs and accelerate degradation of numerous pharmaceuticals (Hicks et al. 2017), but both oxidation and denitrification (N removal) can increase emissions of N 2 O, a greenhouse gas and major ozonedepleting substance (Kampschreur et al. 2009, Ravishankara et al. 2009, Law et al. 2012). On the other hand, adding denitrification to remove N may provide benefits that outweigh cHAB mitigation, such as lower production of nitrogenous cyanotoxins (Downing et al. 2005, Orihel et al. 2012, Horst et al. 2014, Gobler et al. 2016, smaller risk of toxic levels of ammonia and nitrate in surface waters, and improvement of hypoxia in marine coastal zones (Glibert et al. 2006, Paerl 2009). ...
Article
Molot LA, Schiff SL, Venkiteswaran JJ, Baulch HM, Higgins SN, Zastepa A, Verschoor MJ, Walters D. 2021. Low sediment redox promotes cyanobacteria blooms across a trophic range: implications for management. Lake Reserv Manage. XX:XXX-XXX. Field observations and experimental manipulations with different oxidizing agents including nitrate demonstrate that high sediment redox prevents cyanobacteria blooms in eutrophic freshwaters. Conversely, low sediment redox caused by depletion of dissolved oxygen and nitrate allows blooms to form. This explains why bloom risk increases with phosphorus levels: Higher productivity increases the spatial and temporal extent of low sediment redox. The intermediate link between low redox and cyanobacteria blooms appears to be internal loading of ferrous iron (Fe²⁺) from reduced sediments with diffusion to depths accessible to migrating cyanobacteria, providing a source for their high iron demand. Regardless of whether Fe²⁺ release is the intermediate link, the concept of “low sediment redox as promoter” has major potential to improve bloom management if managers consider the impact of their nutrient management choices, nutrient targets, and in-lake methods on sediment redox. Phosphorus input targets can be adjusted as climate change alters the extent of anoxia, and short-term bloom prediction models that incorporate the sediment redox concept could predict onset of blooms earlier than current models that depend on detection of photosynthetic pigments associated with blooms.
... The alarming increase in the detection of ovotestis, testiova, imposex and testicular oocyte conditions are observed among gonochoristic teleosts inhabiting chemically contaminated sites of the aquatic environment (Table 2). Furthermore, reports of intersexuality show either feminization (oocytes within testicular tissue) or masculinization (spermatogenic cells within the ovarian tissue) conditions among teleost fishes (Sardi et al. 2015a;Grilo and Rosa 2017;Hicks et al. 2017). The effects of intersexuality resulting from exposure to xenoestrogenic contaminants include delayed spermatogenesis and oogenesis in males and females, respectively (Bahamonde et al. 2015). ...
Article
Xenoestrogens mimic and interfere with natural functions of oestrogens and adversely affect fish reproduction. Pesticides, plastics, wastewaters and pharmaceuticals are sources of xenoestrogens, and are carried through surface runoffs to water bodies at concentration levels that are harmful to aquatic organisms. Fish absorb xenoestrogens through ingestion of contaminated food items, respiratory gills and dermal contact. Xenoestrogens bioaccumulate in fish tissues, eliciting various reproductive abnormalities, e.g. males may abnormally produce vitellogenins and present with reduced sperm counts, whereas females experience reduced fecundity and hatchability of eggs. Through the food web, xenoestrogens biomagnify in fish predators, e.g. seals and humans, which in turn risk suffering from reproductive malfunctions. Studies of adverse impact of xenoestrogens on fish have mainly been limited to developed countries, yet fish are a major food and trade commodity for developing sub-Saharan African countries. This review serves as a basis for research on adverse impacts of xenoestrogens on fish reproduction, and other consumers of aquatic organisms in Lake Victoria. The lake receives high levels of pollutants from untreated or poorly-treated domestic and industrial wastes and agro-chemicals. Control of xenoestrogens requires concerted effort from multistakeholders to undertake activities such as surveillance, advocacy, legislation and biodegradation to minimise their adverse impacts.
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Designing technologies that mitigate the low-dose adverse effects of exposures to large-volume everyday-everywhere chemicals such as bisphenol A (BPA, 1a ) requires an understanding of the scope of the exposures and...
Article
The present study examined in vitro 11-ketotestosterone and testosterone production by the testes of rainbow darter (Etheostoma caeruleum) collected from selected reference sites and downstream of two municipal wastewater treatment plants (MWWTPs, Waterloo and Kitchener) in the central Grand River (Ontario, Canada), over six years (2011 - 2016). The main objective was to investigate if infrastructure upgrades at the Kitchener MWWTP in 2012 resulted in a recovery of this response in the post-upgrade period (2013 - 2016). Two supporting studies demonstrated that the fall season is appropriate for measuring in vitro sex steroid production as it provides stable detection of steroid patterns, and that the sample handling practiced in this study did not introduce a bias. Infrastructure upgrades of the Kitchener MWWTP resulted in significant reductions in ammonia and estrogenicity. Following the upgrades, 11-ketotestosterone production by MWWTP-exposed fish increased in 2013 and it continued to recover throughout the study period 2014-2016 returning to levels measured in reference fish. Testosterone production was less sensitive and it lacked consistency. Waterloo MWWTP underwent some minor upgrades but the level of ammonia and estrogenicity remained variable over time. The production of 11-ketotestosterone and testosterone in rainbow darter below the Waterloo MWWTP was variable and without a clear recovery pattern over the course of the study. The results of the present study demonstrated that measuring production of sex steroids (especially 11-ketotestosterone) over multiple years can be relevant for assessing responses in fish to environmental changes such as those resulting from major infrastructure upgrades. This article is protected by copyright. All rights reserved.
Article
Decades of studies on endocrine disruption have suggested the need to manage the release of key estrogens from municipal wastewater treatment plants (WWTP). However, the proposed thresholds are below the detection limits of most routine chemical analysis, thereby restricting the ability of watershed managers to assess the environmental exposure appropriately. In this study, we demonstrated the utility of a mechanistic model to address the data gaps on estrogen exposure. Concentrations of the prominent estrogenic contaminants in wastewaters (estrone, estradiol, and ethinylestradiol) were simulated in the Grand River in southern Ontario (Canada) for nine years, including a period when major WWTP upgrades occurred. The predicted concentrations expressed as total estrogenicity (E2 equivalent concentrations) were contrasted to a key estrogenic response (i.e., intersex) in rainbow darter (Etheostoma caeruleum), a wild sentinel fish species. A predicted total estrogenicity in the river of ≥10 ng/L E2 equivalents was associated with high intersex incidence and severity, whereas concentrations <0.1 ng/L E2 equivalents were associated with minimal intersex expression. Exposure to a predicted river concentration of 0.4 ng/L E2 equivalents, the environmental quality standard (EQS) proposed by the European Union for estradiol, was associated with 34% (95% CI:30-38) intersex incidence and a very low severity score of 0.6 (95% CI:0.5-0.7). This exposure is not predicted to cause adverse effects in rainbow darter. The analyses completed in this study were only based on the predicted presence of three major estrogens (E1, E2, EE2), so caution must be exercised when interpreting the results. Nevertheless, this study illustrates the use of models for exposure assessment, especially when measured data are not available.
Article
Gut microbial communities are vital for maintaining host health, and are sensitive to diet, environment, and chemical exposures. Wastewater treatment plants (WWTPs) release effluents containing antimicrobials, pharmaceuticals, and other contaminants that may negatively affect the gut microbiome of downstream organisms. This study investigated changes in the diversity and composition of the digestive gland microbiome of flutedshell mussels (Lasmigona costata) from upstream and downstream of two large (service >100,000) WWTPs. Mussel digestive gland microbiome was analyzed following the extraction, PCR amplification, and sequencing of bacterial DNA using the V3-V4 hypervariable regions of the 16 S rRNA gene. Bacterial alpha diversity decreased at sites downstream of the second WWTP and these sites were dissimilar in beta diversity from sites upstream and downstream of the first upstream WWTP. The microbiomes of mussels collected downstream of the first WWTP had increased relative abundances of Actinobacteria, Bacteroidetes, and Firmicutes, with a decrease in Cyanobacteria, compared to upstream mussels. Meanwhile, those collected downstream of the second WWTP increased in Proteobacteria and decreased in Actinobacteria, Bacteroidetes, and Tenericutes. Increased Proteobacteria has been linked to adverse effects in mammals, but their functions in mussels is currently unknown. Finally, effluent-derived bacteria were found in the microbiome of mussels downstream of both WWTPs but not in those from upstream. Overall, results show that the digestive gland microbiome of mussels collected upstream and downstream of WWTPs differed, which has implications for altered host health and the transport of WWTP-derived bacteria through aquatic ecosystems.
Article
Endocrine active compounds (EACs) are pollutants that have been recognized as an emerging and widespread threat to aquatic ecosystems globally. Intersex, the presence of female germ cells within a predominantly male gonad, is considered a biomarker of endocrine disruption caused by EACs. We measured a suite of EACs and assessed their associated impacts on fish intersex occurrence and severity in a large, regulated river system in North Carolina and South Carolina, USA. Our specific objective was to determine the relationship of contaminants in water, sediment, and fish tissue with the occurrence and severity of the intersex condition in wild, adult black bass (Micropterus), sunfish (Lepomis), and catfish (Ictaluridae) species at 11 sites located on the Yadkin-Pee Dee River. Polycyclic aromatic hydrocarbons (PAHs), ethinylestradiol (EE2), and heavy metals were the most prevalent contaminants that exceeded effect levels for the protection of aquatic organisms. Fish intersex condition was most frequently observed and most severe in black basses and was less frequently detected and less severe in sunfishes and catfishes. The occurrence of the intersex condition in fish showed site-related effects, rather than increasing longitudinal trends from upstream to downstream. Mean black bass and catfish tissue contaminant concentrations were higher than that of sunfish, likely because of the latter's lower trophic position in the food web. Principal component analysis identified waterborne PAHs as the most correlated environmental contaminant with intersex occurrence and severity in black bass and sunfish. As indicated by the intersex condition, EACs have adverse but often variable effects on the health of wild sport fishes in this river, likely due to fluctuations in EAC inputs and the dynamic nature of the riverine system. These findings enhance the understanding of the relationship between contaminants and fish health and provide information to guide ecologically comprehensive conservation and management decisions.
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Recent evidence has revealed that cities with pharmaceutical manufacturers have elevated concentrations of Active Pharmaceutical Ingredients (APIs) in their receiving water bodies. The purpose of this study was to gather information on direct sewer discharges of APIs during their manufacturing and processing from five pharmaceutical manufacturing facilities in Ontario, Canada. Drug classes and maximum reported concentrations (ng/L) for which APIs were directly discharged included: antidepressants (paroxetine - 3380 and sertraline - 5100); mood stabilizer (carbamazepine - 575,000); antibiotics (penicillin - 14,300); analgesics (acetaminophen - 461,000; codeine - 49,200; ibuprofen - 344,000; naproxen - 253,000 and oxycodone 21,000); cardiovascular drugs (atorvastatin - 893 and metoprolol - 7,333,600) and those drugs used for blood pressure control (amlodipine - 22,900; diltiazem - 1,160,000; furosemide - 1,200,000 and verapamil - 7340). Based on flow and water usage data from the individual facilities, the maximum concentrations for acetaminophen, ibuprofen, carbamazepine, diltiazem and metoprolol correlate to approximately 200, 220, 390, 420 and 14,200 g respectively, of lost product being directly discharged to the sewers daily during active manufacturing. This survey demonstrates that direct point source discharges from pharmaceutical manufacturers represent a key source of pharmaceutical pollution to receiving sewersheds. Onsite recovery of product or treatment at pharmaceutical manufacturing or processing facilities to reduce the sewage loadings to receiving treatment plants, product loss and potential environmental loadings is strongly recommended.
Chapter
In our current ecological moment, our planet is becoming increasingly toxic as a result of human consumption. Within this toxic era, media outlets, scientists, and environmentalists rely on intersex human and non-human animal bodies to make obvious the negative effects of human made chemicals on our Planet. Malin Ah-King and Eva Hayward argue that instead of engaging in this “transsex panic” we must understand sex and gender as part of an ongoing process of sexing. Ah-King and Hayward’s understanding of sex as an ever-evolving process does not, however, contend with intersex human bodies that are bearing the physical burden of “transsex panic” through immediate medical intervention at birth and beyond. In response, this chapter offers a reading of Aaron Apps’ poetry collection entitled Intersex: A Memoir, as a text that imagines the speaker’s intersex body as already toxified and magnificent so as to represent intersex thriving in our toxic age.
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During recent decades, survey studies have documented the widespread presence of oocytes in the testes of male Smallmouth Bass Micropterus dolomieu collected from surface waters throughout the United States. There are few published reports of testicular oocytes (TO) in Smallmouth Bass before the 1990s, so it is difficult to know how long this has been occurring. Consequently, this study was conducted to evaluate the prevalence and severity of TO occurrence in whole fish specimens from two archival collections-the Smithsonian Institution's National Museum of Natural History in Suitland, Maryland, and Cornell University's Museum of Vertebrates in Ithaca, New York. Gonads were excised from 167 preserved male Smallmouth Bass that were originally collected between 1875 and 2004, and routine histologic sections were prepared and examined. The severity of TO was determined using a semiquantitative scoring system. Overall, 52.1% of male Smallmouth Bass were found to have TO. Affected fish had been collected in 11 of the 18 represented states, and TO were found in specimens harvested during decades as early as the 1880s and 1900s. Unfortunately, the small number of samples acquired at the earliest time periods precluded analyses of prevalence and severity trends over time. The results of this study demonstrated that the phenomenon of TO in male Smallmouth Bass is at least a century old and confirmed the widespread nature of this finding throughout the species' historic range. Further research efforts should focus on determining the baseline prevalence of TO in laboratory-reared male Smallmouth Bass that have not been exposed to endocrine active substances or the effects of experimental estrogen exposure on such fish.
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Signs of disease, such as external lesions, have been prevalent in smallmouth bass throughout the Susquehanna River Basin, USA. Previous targeted chemical studies in this system have identified known persistent organic pollutants, but a common explanatory link across multiple affected sites remains undetermined. A fast and robust extraction method that can be applied to young-of-year fish is needed to effectively screen for target and non-target compounds that may be impacting organism health. The quick, easy, cheap, effective, rugged, and safe (QuEChERS) extraction methodology was optimized to perform both targeted and non-targeted chemical analyses from a single extraction of whole young-of-year fish. Comprehensive two-dimensional gas chromatography coupled with time-of-flight mass spectrometry (GC×GC-TOFMS) was used for extract analysis. Sample extraction was performed using the solvent ethyl acetate, followed by a two-step cleanup in which samples were frozen for lipid removal and subjected to dispersive solid phase extraction using Florisil. A sample of 21 young-of-year smallmouth bass collected from areas with disease and exhibiting different types of external lesions were evaluated for 233 target compounds. A total of 34 organic contaminants, including polychlorinated biphenyls, brominated diphenyl ethers, organochlorinated pesticides, and personal care products, were detected. Data from this sample set was then analyzed for non-targets. Using the Fisher ratio method and multivariate analysis, an additional 10 significant features were identified specific to either fish with visible lesions or with no visible disease characteristics.
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Effluents from municipal wastewater treatment plants (MWTPs) are complex mixtures of chemicals including endocrine-disrupting compounds (EDCs) and 17α-ethynylestradiol (EE2). The objective of this study was to evaluate selected responses of two fish species, in two different years, exposed in situ to MWTP effluent. Biological markers of exposure (plasma vitellogenin (VTG) and antioxidant enzymes) were measured in two species of male fish, rainbow trout (Oncorhynchus mykiss) and fathead minnow (Pimephales promelas), caged at sites associated with wastewater outfall. The estrogenicity of the final effluent in 2010 was determined to be 17.0 + 0.4 ng/L estrogen equivalents (EEQ) and reduced to 7.5 + 2.9 ng/L EEQ after infrastructure upgrades. Pharmaceuticals and personal care products in the effluent and surface water in both years confirmed the exposures at each downstream site. Despite the presence of estrogenic compounds in the MWTP effluent, no effluent-caged male fish demonstrated plasma VTG induction. Minnows and trout that received an intraperitoneal injection of 5 mg/g EE2 showed VTG induction at both field sites. In 2012, the liver somatic index (LSI) of both species increased with exposure, as did changes in antioxidant enzymes, and reactive oxygen species (ROS) activity. Multiple biological mechanisms are modified by effluent exposure, and multiple endpoints are needed to assess risk. HIGHLIGHTS Effluent exposure resulted in induction of antioxidant enzymes and ROS activity in caged fish.; Despite measurable estrogen equivalents in the effluent, downstream caged male fathead minnow did not demonstrate VTG induction.; EE2 injected, control male fish exposed to wastewater or upstream reference conditions responded with elevated VTG concentrations.; Several factors may have reduced the VTG response in caged fish.;
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The application of some hormones in fish production in swamps enhances fish production. However, in this regard, the critical issue is chemical contamination in water, which affects aquatic organisms. Steroid hormones (e.g., androgens and estrogens) can change the sexual characteristics of aquatic organisms, especially fish. These compounds are used in oral contraceptives and have an effect on fish at different developmental stages, such as the development of an embryo. The residual effects of steroid hormones can adversely affect the environment, biological activities, and human health. Furthermore, these hormones negatively affect the endocrine system in marine organisms. Therefore, the identification and measure of these compounds by sensitive and selective analytical methods is not only essential, but can also prevent their adverse effects. Estrone (E1), 17β-estradiol (17β-E2), and 17α-ethynylestradiol (EE2) are current steroid hormones, which create feminized properties in male fish. The concentration of E1 in aquatic surroundings is more than the concentration of estradiol (E2) and EE2; however, E1 has a low connection and activation power to the estrogen receptor (ER). Hence, the 17β-hydroxysteroid dehydrogenases (17β-HSDs) enzyme converts E1 to E2, especially in an environment with a high E1 concentration, such as wastewater treatment. Researchers showed that E1 and its conversion have dangerous impacts on the generation and breeding of different fish species in polluted aquatic environments. In this regard, the goal of this systematic review is to evaluate the role of steroid hormones in aquatic organisms, especially fish, as well as to evaluate the negative consequences of these compounds on humans and the surrounding.
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Municipal wastewater treatment plants (WWTPs) contain numerous contaminants, including antimicrobials, that could affect the composition of the beneficial bacterial communities associated with host aquatic organisms. There is also potential for these effects to transfer to terrestrial predators. Riparian spiders and five families of aquatic macroinvertebrates were collected from sites upstream and downstream of two WWTPs, Waterloo and Kitchener, discharging to the Grand River, Ontario, Canada. Whole-body microbiota were analyzed following the extraction, PCR amplification, and sequencing of bacterial DNA using the V3-V4 hypervariable regions of the 16S rRNA genetic barcode. Changes in the relative abundance of major microbiome phyla were observed in all downstream aquatic insects except Hydropsychidae caddisflies, which exhibited little site variation. Shannon alpha diversity differed among sites for Tetragnathidae spiders, Perlidae, Hydropsychidae, and Heptageniidae. Downstream of the Waterloo WWTP alpha diversity decreased in spiders, while downstream of the Kitchener WWTP this measure decreased in Perlidae and increased in spiders. Bray-Curtis beta diversity was dissimilar among sites in all invertebrate taxa; upstream sites differed from those downstream of Waterloo in spiders, Perlidae, and Hydropsychidae, and from those downstream of Kitchener in spiders, Perlidae, and Hydropsychidae. Finally, effluent-derived bacteria were found in the microbiomes of downstream spiders and aquatic insects and not upstream. Overall, results indicated that the microbiomes of invertebrates collected downstream differed from those collected upstream of WWTPs, which has implications for altered host health and transport of WWTP-derived bacteria through aquatic ecosystems.
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While wastewater treatment standards have been progressively increasing, emerging contaminants such as pharmaceuticals can nonetheless pass through treatment and end up in our watersheds. Pharmaceuticals in the parts-per-billion range can impact fish behavior, survival, and recruitment in the wild. However, the ecological risk posed by whole municipal wastewater effluents (MWWE), a complex mixture, is not clear. This knowledge gap is particularly evident for early lifestages (ELS) of fish, and because effluent discharge events are typically short, the effects of short-term MWWE exposures to ELS fish are particularly important from an environmental perspective. Here we tested the effects of rapid 30-min exposures, and short-term 24- and 72-h exposures to MWWE on development, behaviors, and stress response in zebrafish (Danio rerio) embryos, larvae, and juveniles. We obtained 24-h composite samples of tertiary-treated MWWE that contained a mixture of chemicals with affinities for serotonergic, adrenergic, dopaminergic, and ion-channel receptors. Embryos exposed to 5%, 10%, and 50% MWWE experienced developmental delays in somitogenesis and hatching rate, although there was no effect on survival. Embryonic photomotor responses were affected following 30-min and 24-h exposures to 10% and 50% MWWE, and larval visual motor responses were reduced from 24-h exposure to 10% MWWE. Exposure to 10% MWWE dulled the juvenile cortisol and lactate response following an acute air-exposure. Compromised behavioral and stress performances demonstrate the capacity of MWWE to impact phenotypes critical to the survival of fish in the environment. Taken together, we found that zebrafish were sensitive to toxic effects of MWWE at multiple life-stages.
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Municipal wastewater effluent (MWWE) and its constituents, such as chemicals of emerging concern, pose a potential threat to the sustainability of fish populations by disrupting key endocrine functions in aquatic organisms. While studies have demonstrated changes in biological markers of exposure of aquatic organisms to groups of chemicals of emerging concern, the variability of these markers over time has not been sufficiently described in wild fish species. The aim of this study was to assess the spatial and temporal variability of biological markers in response to MWWE exposure and to test the consistency of these responses between seasons and among years. Rainbow darter ( Etheostoma caeruleum ) were collected in spring and fall seasons over a 5-year period in the Grand River, Ontario, Canada. In addition to surface water chemistry (nutrients and selected pharmaceuticals), measures were taken across levels of biological organization in rainbow darter. The measurements of hormone production, gonad development, and intersex severity were temporally consistent and suggested impaired reproduction in male fish collected downstream of MWWE outfalls. In contrast, ovarian development and hormone production in females appeared to be influenced more by urbanization than MWWE. Measures of gene expression and somatic indices were highly variable between sites and years, respectively, and were inconclusive in terms of the impacts of MWWE overall. Robust biomonitoring programs must consider these factors in both the design and interpretation of results, especially when spatial and temporal sampling of biological endpoints is limited. Assessing the effects of contaminants and other stressors on fish in watersheds would be greatly enhanced by an approach that considers natural variability in the endpoints being measured.
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The occurrence of intersex fish, where male reproductive tissues show evidence of feminization, have been found in freshwater systems around the world, indicating the potential for significant endocrine disruption across species in the ecosystem. Estrogens from birth control medications in wastewater treatment plant effluent have been cited as the likely cause, but research has shown that endocrine disruption is not solely predictable based on hormone receptor interactions. Many other non-hormone pharmaceuticals are found in effluent at concentrations orders of magnitude higher than estrogens, yet there is little data indicating the impacts of these other medications. The widely prescribed anti-diabetic metformin is among the most abundant of pharmaceuticals found in effluent and is structurally dissimilar from hormones. However, we show here that exposing fathead minnows (Pimephales promelas) to a concentration of metformin found in wastewater effluent causes the development of intersex gonads in males, reduced size of treated male fish, and reduction in fecundity for treated pairs. Our results demonstrate that metformin acts as an endocrine disruptor at environmentally relevant concentrations. Copyright © 2015 The Authors. Published by Elsevier Ltd.. All rights reserved.
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Endocrine-disrupting chemicals (EDCs) in municipal effluents directly affect the sexual development and reproductive success of fishes, but indirect effects on invertebrate prey or fish predators through reduced predation or prey availability, respectively, are unknown. At the Experimental Lakes Area in northwestern Ontario, Canada, a long-term, whole-lake experiment was conducted using a before-after-control-impact design to determine both direct and indirect effects of the synthetic oestrogen used in the birth control pill, 17 alpha-ethynyloestradiol (EE2). Algal, microbial, zooplankton and benthic invertebrate communities showed no declines in abundance during three summers of EE2 additions (5-6 ng l(-1)), indicating no direct toxic effects. Recruitment of fathead minnow (Pimephales promelas) failed, leading to a near-extirpation of this species both 2 years during (young-of-year, YOY) and 2 years following (adults and YOY) EE2 additions. Body condition of male lake trout (Salvelinus namaycush) and male and female white sucker (Catostomus commersonii) declined before changes in prey abundance, suggesting direct effects of EE2 on this endpoint. Evidence of indirect effects of EE2 was also observed. Increases in zooplankton, Chaoborus, and emerging insects were observed after 2 or 3 years of EE2 additions, strongly suggesting indirect effects mediated through the reduced abundance of several small-bodied fishes. Biomass of top predator lake trout declined by 23-42% during and after EE2 additions, most probably an indirect effect from the loss of its prey species, the fathead minnow and slimy sculpin (Cottus cognatus). Our results demonstrate that small-scale studies focusing solely on direct effects are likely to underestimate the true environmental impacts of oestrogens in municipal wastewaters and provide further evidence of the value of whole-ecosystem experiments for understanding indirect effects of EDCs and other aquatic stressors.
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1. Increased need for water and projected declines in precipitation due to climate change could leave waterways increasingly dominated by wastewater effluent. Understanding how components of wastewater influence fish populations is necessary for effective conservation and management. Despite research demonstrating effects of oestrogens, such as 17a-ethynylestradiol (EE2), on fish physiology and population failure, the generality of population responses is uncertain and the underlying mechanisms affecting population declines are unknown. EE2 is the steroid oestrogen in human contraceptive pills and has been measured up to 11 ng L ^-1 in the environment. 2. We identify disrupted population dynamics due to direct and transgenerational effects on survival and fecundity. We conducted a year-long study on three generations of fathead minnows Pimephales promelas Rafinesque in aquatic mesocosms and laboratory aquaria. We added environmentally relevant concentrations of EE2 daily using a static renewal, which approximates a pulsed exposure that fish experience in natural systems. 3. EE2 (3*2 ng L ^-1) reduced F0 male survival to 17% (48% lower than controls) and juvenile production by 40% compared to controls. F1 fish continuously exposed to EE2 failed to reproduce, and reproduction of the F1 transferred to clean water was 70–99% less than controls. 4. F2 larval survival, exposed only as germ cells in their parents, was reduced by 51–97% compared to controls. The indirect effect on F2 survival suggests the possibility of transgenerational effects of EE2. 5. Synthesis and applications. Our results suggest that fish populations exposed to environmentally relevant 17a-ethynylestradiol (EE2) concentrations may not recover from exposure. Management of short-lived highly fecund fishes should be prioritized to protect fish from the embryo through gonadal differentiation. Reducing effluent will not be possible in many situations; hence, conservation of breeding and rearing habitat in unpolluted tributaries or reaches is needed. Additionally, resource managers could enhance habitat connectivity in rivers to facilitate immigration. Finally, investment in advanced wastewater processing technology should improve removal of bioactive chemicals such as EE2. Our results provide a baseline for regulatory agencies to consider when assessing the ecological effects of environmental oestrogens, and our approach to evaluating population-level effects could be widely applied to other contaminants.
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The variability and extent of the intersex condition (oocytes in testes or testis-ova) in fish along an urban gradient, that included major wastewater treatment plant outfalls, was documented in the Grand River, Ontario, Canada. A rapid enumeration of testis-ova method (RETO) was developed and applied that increased the capacity to quantify both intersex prevalence and severity. Male Rainbow Darters (Etheostoma caeruleum) sampled downstream of the first major wastewater outfall (Waterloo) had a significant increase, relative to four upstream reference sites, in the mean proportion of fish with at least one testis-oocyte (t.o.) per lobe of testes (9-20% proportion with ≤ 1 t.o./lobe vs. 32-53% and >1.4 t.o./lobe). A much higher mean incidence of intersex proportion and degree was observed immediately downstream of the second wastewater outfall (Kitchener; 73-100% and 8-70 t.o./lobe) but only 6.3 km downstream of the Kitchener outfall the occurrence of intersex drops to those of the reference sites. In contrast, downstream of a tertiary treated wastewater outfall on a small tributary intersex was similar to reference sites. Estrogenicity, measured using a Yeast Estrogen Screen (YES), followed a similar pattern increasing from 0.81 ± 0.02 ng/L EEq (Guelph), to 4.32 ± 0.07 ng/L (Waterloo), and 16.99 ± 0.40 ng/L (Kitchener) respectively. Female Rainbow Darter downstream of the Kitchener outfall showed significant decreases in gonadosomatic index (GSI) and liver somatic index (LSI), and increases in condition factor (k) relative to corresponding reference sites. The prevalence of intersex and alterations in somatic indices suggest that exposure to municipal wastewater effluent discharges can impact endocrine function, energy use, and energy storage in wild fish. Environ Toxicol Chem © 2013 SETAC.
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A high prevalence of intersex or testicular oocytes (TO) in male smallmouth bass within the Potomac River drainage has raised concerns as to the health of the river. Studies were conducted to document biomarker responses both temporally and spatially to better understand the influence of normal physiological cycles, as well as water quality and land-use influences. Smallmouth bass were collected over a 2-year period from three tributaries of the Potomac River: the Shenandoah River, the South Branch Potomac and Conococheague Creek, and an out-of-basin reference site on the Gauley River. The prevalence of TO varied seasonally with the lowest prevalence observed in July, post-spawn. Reproductive maturity and/or lack of spawning the previous spring, as well as land-use practices such as application of manure and pesticides, may influence the seasonal observations. Annual, seasonal, and site differences were also observed in the percentage of males with measurable concentrations of plasma vitellogenin, mean concentration of plasma vitellogenin in females, and plasma concentrations of 17β-estradiol and testosterone in both sexes. Bass collected in the South Branch Potomac (moderate to high prevalence of TO) had less sperm per testes mass with a lower percentage of those sperm being motile when compared to those from the Gauley River (low prevalence of TO). An inverse relationship was noted between TO severity and sperm motility. An association between TO severity and wastewater treatment plant flow, percent of agriculture, total number of animal feeding operations, the number of poultry houses, and animal density within the catchment was observed.
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The Grand River watershed in Ontario, Canada, receives and assimilates the outflow of 29 Municipal Wastewater Effluent (MWWE) discharges which is a mixture of domestic and industrial wastes. The purpose of this study was to investigate the cumulative impact of multiple sewage discharges on populations of wild fish. In field studies, responses of fish populations and individual fish responses in terms of growth (condition factor), reproduction (in vitro sex steroid production, gonadosomatic indices, histology [cellular development and intersex]) were assessed upstream and downstream of two municipal discharges. Fish [Greenside Darters Etheostoma blennioides and Rainbow Darters E. caeruleum] collected downstream of two municipal wastewater plants had the potential to have greater growth (longer and heavier) when compared to reference fish collections regardless of sex. Fish were not assimilating additional anthropogenic resources into energy storage (increased condition, liver somatic index). Impacts on ovarian development appeared to be minor with no differences in growth, steroid production or cellular development. Sewage exposed male fish were experiencing impairment in the capacity to produce testosterone and 11-ketotestosterone in vitro, and in cellular development (GSI, intersex). Male darters of both species collected in the upstream agricultural region demonstrated no evidence of intersex whereas our urban reference sites had incidence of intersex of up to 20%. Rates of intersex were elevated downstream of both sewage discharges studied (33% and>60%, respectively). Lower rates of intersex at the intermediate sites, and then increases downstream of second sewage discharge suggests that fish populations have to potential to recover prior to exposure to the second sewage effluent. Pre-spawning darters demonstrated dramatically higher incidence of intersex in the spring at both urban reference sites (33% and 50%, respectively), and increased more so downstream of the near-field and far-field exposure sites (60% and 100%, respectively). These findings suggest that the compounds released in STP effluents have a tendency to act on the male reproductive system. These effects may become more pronounced as projected human population growth will require the aquatic environment to assimilate an increasing amount of sewage waste.
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The feminization of nature by endocrine-disrupting chemicals (EDCs) is a key environmental issue affecting both terrestrial and aquatic wildlife. A crucial and as yet unanswered question is whether EDCs have adverse impacts on the sustainability of wildlife populations. There is widespread concern that intersex fish are reproductively compromised, with potential population-level consequences. However, to date, only in vitro sperm quality data are available in support of this hypothesis. The aim of this study was to examine whether wild endocrine-disrupted fish can compete successfully in a realistic breeding scenario. In two competitive breeding experiments using wild roach (Rutilus rutilus), we used DNA microsatellites to assign parentage and thus determine reproductive success of the adults. In both studies, the majority of intersex fish were able to breed, albeit with varying degrees of success. In the first study, where most intersex fish were only mildly feminized, body length was the only factor correlated with reproductive success. In the second study, which included a higher number of more severely intersex fish, reproductive performance was negatively correlated with severity of intersex. The intersex condition reduced reproductive performance by up to 76% for the most feminized individuals in this study, demonstrating a significant adverse effect of intersex on reproductive performance. Feminization of male fish is likely to be an important determinant of reproductive performance in rivers where there is a high prevalence of moderately to severely feminized males.
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The yeast estrogen screen was applied to sewage treatment process waters to identify the presence of estrogenic activity and to investigate the fate and behavior of estrogenic substances through treatment. Hydrophobic fractions in the water phase were extracted and concentrated using C18 cartridges for the effective extraction of 17β-estradiol (E2) and other estrogen mimics. Clear dose-dependent elevation in the synthesis of β-galactosidase in the yeast screen was observed with all the samples tested, demonstrating that these samples were estrogenic. However, estrogenic activity tended to reduce during the treatment, especially in the biological process. Quantification results of the yeast estrogen screen in terms of E2 equivalent were compared with actual E2 concentrations measured by an ELISA. E2 occupied 34% of the whole estrogenicity in the raw sewage, while almost 100% in the final effluent. The analyses of the sewage treatment process waters revealed that human estrogens are major causative substances in terms of estrogenic activity in sewage and its treated effluent. Although findings of possible correlation of environmental estrogens with the real impact on human health and the ecosystem are still the focus of scientific debate and investigation, proper management should be established in the sewage treatment process which receives various environmental contaminants.
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The widespread occurrence of feminized male fish downstream of some wastewater treatment works has led to substantial interest from ecologists and public health professionals. This concern stems from the view that the effects observed have a parallel in humans, and that both phenomena are caused by exposure to mixtures of contaminants that interfere with reproductive development. The evidence for a "wildlife-human connection" is, however, weak: Testicular dysgenesis syndrome, seen in human males, is most easily reproduced in rodent models by exposure to mixtures of antiandrogenic chemicals. In contrast, the accepted explanation for feminization of wild male fish is that it results mainly from exposure to steroidal estrogens originating primarily from human excretion. We sought to further explore the hypothesis that endocrine disruption in fish is multicausal, resulting from exposure to mixtures of chemicals with both estrogenic and antiandrogenic properties. We used hierarchical generalized linear and generalized additive statistical modeling to explore the associations between modeled concentrations and activities of estrogenic and antiandrogenic chemicals in 30 U.K. rivers and feminized responses seen in wild fish living in these rivers. In addition to the estrogenic substances, antiandrogenic activity was prevalent in almost all treated sewage effluents tested. Further, the results of the modeling demonstrated that feminizing effects in wild fish could be best modeled as a function of their predicted exposure to both antiandrogens and estrogens or to antiandrogens alone. The results provide a strong argument for a multicausal etiology of widespread feminization of wild fish in U.K. rivers involving contributions from both steroidal estrogens and xenoestrogens and from other (as yet unknown) contaminants with antiandrogenic properties. These results may add further credence to the hypothesis that endocrine-disrupting effects seen in wild fish and in humans are caused by similar combinations of endocrine-disrupting chemical cocktails.
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Endocrine-disrupting chemicals, known to be present in the environment, have great potential for interfering with reproductive health in wildlife and humans. There is, however, little direct evidence that endocrine disruption has adversely affected fertility in any organism. In freshwater and estuarine fish species, for example, although a widespread incidence of intersex has been reported, it is not yet known if intersexuality influences reproductive success. The purpose of this study was, therefore, to determine gamete quality in wild intersex roach (Rutilus rutilus) by assessing sperm characteristics, fertilization success, and ability to produce viable offspring. The results clearly demonstrate that gamete production is reduced in intersex roach. A significantly lower proportion of moderately or severely feminized fish (17.4% and 33.3%, respectively) were able to release milt compared with normal male fish from contaminated rivers (in which 97.6% of the males were able to release milt), reference male fish (97.7%), or less severely feminized intersex fish (experiment 1: 85.8%, experiment 2: 97%). Intersex fish that did produce milt produced up to 50% less (in terms of volume per gram of testis weight) than did histologically normal male fish. Moreover, sperm motility (percentage of motile sperm and curvilinear velocity) and the ability of sperm to successfully fertilize eggs and produce viable offspring were all reduced in intersex fish compared with normal male fish. Male gamete quality (assessed using sperm motility, sperm density, and fertilization success) was negatively correlated with the degree of feminization in intersex fish (r = -0.603; P < 0.001) and was markedly reduced in severely feminized intersex fish by as much as 50% in terms of motility and 75% in terms of fertilization success when compared with either less severely feminized intersex fish or unaffected male fish. This is the first evidence documenting a relationship between the morphological effects (e.g., intersex) of endocrine disruption and the reproductive capabilities of any wild vertebrate. The results suggest that mixtures of endocrine-disrupting substances discharged into the aquatic environment could pose a threat to male reproductive health.
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Surveys of U.K. rivers have shown a high incidence of sexual disruption in populations of wild roach (Rutilus rutilus) living downstream from wastewater treatment works (WwTW), and the degree of intersex (gonads containing both male and female structural characteristics) has been correlated with the concentration of effluent in those rivers. In this study, we investigated feminized responses to two estrogenic WwTWs in roach exposed for periods during life stages of germ cell division (early life and the postspawning period). Roach were exposed as embryos from fertilization up to 300 days posthatch (dph; to include the period of gonadal sex differentiation) or as postspawning adult males, and including fish that had received previous estrogen exposure, for either 60 or 120 days when the annual event of germ cell proliferation occurs. Both effluents induced vitellogenin synthesis in both life stages studied, and the magnitude of the vitellogenic responses paralleled the effluent content of steroid estrogens. Feminization of the reproductive ducts occurred in male fish in a concentration-dependent manner when the exposure occurred during early life, but we found no effects on the reproductive ducts in adult males. Depuration studies (maintenance of fish in clean water after exposure to WwTW effluent) confirmed that the feminization of the reproductive duct was permanent. We found no evidence of ovotestis development in fish that had no previous estrogen exposure for any of the treatments. In wild adult roach that had previously received exposure to estrogen and were intersex, the degree of intersex increased during the study period, but this was not related to the immediate effluent exposure, suggesting a previously determined programming of ovotestis formation.
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A number of chemicals present in the environment have been shown to mimic or antagonize the actions of steroid hormones, an issue often described as "endocrine disruption/modulation". There is very little evidence, however, to support the hypothesis that exposure to endocrine-disrupting chemicals is a global environmental health problem. In this paper, we demonstrate a high incidence of intersexuality in wild populations of riverine fish (roach; Rutilus rutilus) throughout the United Kingdom. These reproductive disturbances are consistent with exposure to hormonally active substances and are associated with discharges from sewage treatment works that are known to contain estrogenic chemicals. This is the first documented example of a widespread sexual disruption in wild populations of any vertebrate and indicates that reproductive and developmental effects do result from exposure to ambient levels of chemicals present in typical British rivers.
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